Dominant-Negative Calcium Channel Suppression by Truncated Constructs Involves a Kinase Implicated in the Unfolded Protein Response

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Abstract
Expression of the calcium channel CaV2.2 is markedly suppressed by coexpression with truncated constructs of CaV2.2. Furthermore, a two-domain construct of CaV2.1 mimicking an episodic ataxia-2 mutation strongly inhibited CaV2.1 currents. We have now determined the specificity of this effect, identified a potential mechanism, and have shown that such constructs also inhibit endogenous calcium currents when transfected into neuronal cell lines. Suppression of calcium channel expression requires interaction between truncated and full-length channels, because there is inter-subfamily specificity. Although there is marked cross-suppression within the CaV2 calcium channel family, there is no cross-suppression between CaV2 and CaV3 channels. The mechanism involves activation of a component of the unfolded protein response, the endoplasmic reticulum resident RNA-dependent kinase (PERK), because it is inhibited by expression of dominant-negative constructs of this kinase. Activation of PERK has been shown previously to cause translational arrest, which has the potential to result in a generalized effect on protein synthesis. In agreement with this, coexpression of the truncated domain I of CaV2.2, together with full-length CaV2.2, reduced the level not only of CaV2.2 protein but also the coexpressed α2δ-2. Thapsigargin, which globally activates the unfolded protein response, very markedly suppressed CaV2.2 currents and also reduced the expression level of both CaV2.2 and α2δ-2 protein. We propose that voltage-gated calcium channels represent a class of difficult-to-fold transmembrane proteins, in this case misfolding is induced by interaction with a truncated cognate CaV channel. This may represent a mechanism of pathology in episodic ataxia-2.

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